Media formulation

Media formulation for fermentation

1. Dr. N. BANU Vels University

2. Microbe requires Water Energy Carbon Nitrogen Mineral elements Vitamins Oxygen MEDIA FORMULATION

3. Max. yield Max. conc. Of product Min. yield of undesired product Stable product Min. problems during media making and sterilization Min. problems of production processes (aeration, agitation, extraction, purification and waste treatment. Criteria for nutrients for a medium

4. Major component of all fermentation. Ancillary services : heating, cooling, cleaning and rinsing. Suitable water: pH, dissolved salts, effluent contamination. Brewing: mineral content of the water. Reuse of water after fermentation – Methylophilus methylotrophus WATER

5. Carbon &energy source + nitrogen + others STOICHIOMETRY OF GROWTH AND PRODUCT FORMATION BIOMASS + PRODUCTS + CO 2 +H 2 O+ HEAT

6. Oxidation of the medium components (carbohydrates, lipids and proteins, light ENERGY SOURCE

7. Carbohydrates Oils and fats Hydrocarbon and their derivatives Carbohydrates: Starch from maize, cassava, cereals, potatoes Barley grains – malt. Sucrose from sugar cane and sugar beet. Molasses – high volume, low-value product. Lactose and crude lactose Corn steep liquor. Carbon sources

8. First used as carriers for antifoams in antibiotic processes. Vegetable oils – olive, maize, cotton seed, lin seed, soybean, etc. Used as carbon substrates. Also provides antifoaming property. Methly oleate – sole ‘ c’source in cephalosporin production. Oils and fats

9. Methane, methanol and n- alkanes – substrates for biomass production. N- alkanes – for the production of organic acids, amino acids, vitamins and co-factors, nucleic acids, antibiotics, enzymes and proteins. Scp from hydrocarbons: BP plc developed Toprina from yeast grown on n- alkanes . ICI plc developed pruteen from bacteria grown on methanol. Hoechst/UBHE (protein from bacteria on methanol) Shell plc (bacteria on methane). Hydrocarbons and their derivatives

10. Inorganic source and organic source Inorganic Nitrogen: Ammonia gas Ammonium salts Nitrates Organic nitrogen: Amino acids Protein Urea Protenaceous nitrogen compounds: CSL, soya meal, peanut meal, cotton-seed meal, distiller’s solubles , meal and yeast extract. Nitrogen sources

11. Penicillin Bacitracin Riboflavin Novobiocin Rifomycin CSL Peanut granules Pancreatic digest of gelatine Distillers solubles Pharmamedia , soybean meal, (NH 4 ) 2 SO 4. Best N source for some sec. metabolites

12. KH 2 PO 4 MgSO 4 . 7H 2 KCl CaCO 3 FeSo 4 . 4H 2 O ZnSO 4 . 8H 2 MnSo 4 . H 2 O CuSO 4 .5H 2 O Na 2 MoO 4 . 2H 2 1.0-4.0 0.25-3.0 0.5-12.0 5-17 .01-.1 .1-1 .01-.1 .003-.01 .01-.1 Minerals

13. Trace elements influencing primary and secondary metabolism Product Trace elements Bacitracin Protease Gentamicin Riboflavin Chloramphenicol Citric acid Penicillin Patulin Mn Mn Co Fe, Co Fe, Zn Fe, Zn, Cu Fe, Zn, Cu Fe, Zn

14. Vitamins Amino acids, Fatty acids Sterols. Calcium pantothenate – Vinegar production Biotin – glutamic acid production . Growth factors

15. Calcium carbonate Phosphates Balanced use of C/N ratio controls pH pH controlled externally by adding ammonia or sodium hydroxide and sulfuric acid. Buffers

16. Penicillin production – phenlyacetic acid – precursor. Inducers: amylase – starch, maltose. Pullulanase – maltose. Penicillin acylase – phenlyacetic acid Proteases – various proteins Cellulase – cellulose Pectinases - pectin Precursors and inducers

17. Influence of medium for oxygen availability: Fast metabolism: The culture may become oxygen limited because sufficient O2 cannot be made available in the fermentor if certain substrates, like rapidly metabolized sugars which leads to a high oxygen demand, are available in high conc. Rheology : The individual components of the medium can influence the viscosity of the final medium and its subsequent behaviour with respect to aeration and agitation. Antifoams: Act as surface active agents and reduce the o2 transfer rate. Oxygen requirement

18. Foaming due to component of the medium / protein in the medium – leads to problem. Characteristics of antifoams: Should disperse readily and have fast action on an excisting foam. Should be active at low conc. Should be long acting in preventing new foam formation. Should not be metabolized by the microorganisms. Should be non-toxic to the microbe,humans and animals. Should not cause any problems in the downstreaming Should not cause any handling hazards. Should be cheap. Should have no effect on oxygen transfer Should be heat sterilizable . Antifoams

19. Alcohols; stearyl and octyl decanol Esters Fatty acids and derivatives ( glycerides ) linseed oil, cottonseed oil, olive oil, castor oil, sunflower oil, rapeseed oil and cod liver oil. Silicones Sulphonates Alkaterge C, oxazaline , poly-propylene glycol. Antifoams examples